ITK can induce negative regulators that influence the kinetics of CD8+ T cell TCR signaling 3028

Abstract Description The initiation of T cell receptor (TCR) signaling is well characterized and is known to include both positive and negative regulatory factors. The termination of TCR signaling is less well understood. Interleukin-2 inducible T cell kinase (ITK), a member of the TEC family of non-receptor tyrosine kinases, functions to tune the strength of TCR signaling in response to peptides of varying affinities. By utilizing Nr4a3-Tocky reporter mice, we show that the initiation of TCR signaling in ITK KO CD8+ T cells is delayed and of reduced magnitude compared to ITK WT CD8+ T cells. Further, at later timepoints, the downregulation of TCR-induced activation markers and transcription factors is also delayed in the absence of ITK. To address the mechanism(s) underlying ITK-dependent tuning of TCR signaling kinetics, we performed bulk RNA-sequencing on ITK WT and KO CD8+ T cells at 0, 5, 12, and 24 hours after TCR stimulation with high and low affinity peptide ligands. These data revealed that multiple negative regulators of TCR signaling, including Cish, Ptpn11, Dusp12, and Spry1, are poorly upregulated in ITK KO CD8+ T cells after activation compared to ITK WT cells. Currently, studies are in progress to assess the functional relevance of these negative regulators by CRISPR/Cas9 targeting. Overall, our data indicate that ITK functions to induce the upregulation of inhibitory factors that contribute to the termination of TCR signaling. Funding Sources This work was supported by the NIH/NIAID (R01 AI043957). The National Jewish Health Genomics Facility and the Barbara Davis Center Cell and Tissue Analysis Core at the University of Colorado Diabetes Research Center (funded by NIH/NIDDK P30-DK116073) were also utilized. Topic Categories Immune Response Regulation: Molecular Mechanisms (IRM)